Taenia solium cysticercosis/taeniosis is emerging as a serious public health and economic problem in many developing countries. This study was conducted to determine prevalence and risk factors of human T. solium infections in Mbeya Region, Tanzania.
Methods and Findings
A cross-sectional survey was conducted in 13 villages of Mbozi district in 2009. Sera of 830 people (mean 37.9±11.3 years (SD); 43% females) were tested for circulating cysticerci antigen (Ag-ELISA) and antibody (Ab-ELISA). A subset of persons found seropositive by Ag-ELISA underwent computed tomography (CT) scan of the brain for evidence of neurocysticercosis. Stool samples from 820 of the same participants were tested for taeniosis by copro-antigens (copro-Ag-ELISA) and formol-ether concentration technique. Cases of T. solium taeniosis were confirmed serologically by EITB assay (rES38). A questionnaire was used for identification of risk factors. Active cysticercosis by positive Ag-ELISA was found in 139 (16.7%) persons while anti-cysticercal antibodies were detected in 376 (45.3%) persons by Ab-ELISA. Among 55 persons positive for Ag-ELISA undergoing CT scan, 30 (54.6%) were found to have structures in the brain suggestive of neurocysticercosis. Using faecal analysis, 43 (5.2%) stool samples tested positive for taeniosis by copro-Ag-ELISA while Taenia eggs were detected in 9 (1.1%) stool samples by routine coprology. Antibodies specifically against adult T. solium were detected in 34 copro-Ag-ELISA positive participants by EITB (rES38) indicating T. solium taeniosis prevalence of 4.1%. Increasing age and hand washing by dipping in contrast to using running water, were found associated with Ag-ELISA seropositivity by logistic regression. Gender (higher risk in females) and water source were risk factors associated with Ab-ELISA seropositivity. Reported symptoms of chronic severe headaches and history of epileptic seizures were found associated with positive Ag-ELISA (p≤0.05).
The present study indicates T. solium infection in humans is highly endemic in the southern highlands of Tanzania.
Cysticercosis caused by the zoonotic pork tapeworm, Taenia solium, is emerging as a serious public health and agricultural problem in sub-Saharan Africa. Surveys have shown cysticercosis in pigs to be highly prevalent in multiple foci in Tanzania, and a hospital-based study in the northern highlands indicated neurocysticercosis as an important cause of epileptic seizures in humans. We present here a cross-sectional community-based survey on the prevalence and risk factors of human cysticercosis and taeniosis conducted in the southern highlands – the major pig-producing area of the country. The most striking findings were that more than 15% of people surveyed were found to have active cysticercosis and nearly half of them were found to have been exposed to larval T. solium indicating a high level of environmental contamination with T. solium eggs. This was supported by finding over 4% of people having had T. solium tapeworms. A subset of persons found positive serologically for active cysticercosis underwent brain scanning and more than half of them were found to have neurocysticercosis. This strong evidence that T. solium cysticercosis/neurocysticercosis/taeniosis is highly endemic in the southern highlands of Tanzania demands urgent attention of regional and national authorities to combat the parasite.
Src is over-expressed or hyper-activated in a variety of human cancers including thyroid carcinoma. Src is a central mediator in multiple signaling pathways that are important in oncogenesis and cancer progression. In this study, we evaluated the effects of a Src inhibitor, SKI-606 (bosutinib), in a spontaneous metastatic thyroid cancer model with constitutively activated Src (ThrbPV/PVPten+/− mice).
ThrbPV/PVPten+/− mice were treated with SKI-606 or vehicle controls, beginning at 6 weeks of age until the mice succumbed to thyroid cancer. We assessed the effects of SKI-606 on thyroid cancer progression and analyzed the impact of SKI-606 on aberrant Src-mediated signaling.
SKI-606 effectively inhibited aberrant activation of Src and its downstream targets to markedly inhibit the growth of thyroid tumor, thereby prolonging the survival of treated mice. While Src inhibition did not induce cell apoptosis, it decreased cell proliferation by affecting the expression of key regulators of cell cycle progression. Importantly, SKI-606 dramatically prevented de-differentiation, vascular invasion, and lung metastasis of thyroid cancer cells. These responses were meditated by down-regulation of mitogen-activated protein kinase pathways and inhibition of the epithelial-mesenchymal transition.
Our findings suggest that Src is critical in the progression of thyroid cancer, making oral SKI-606 a promising treatment strategy for refractory thyroid cancer.
Thyroid cancer; Src; SKI-606; metastasis; epithelial-mesenchymal transition; mouse model
Hereditary pancreatitis shares a majority of clinical and morphologic features with chronic alcoholic pancreatitis, but may present at an earlier age. The term hereditary pancreatitis has primarily been associated with mutations in the serine protease 1 gene (PRSS1) which encodes for cationic trypsinogen. PRSS1 mutations account for approximately 68–81% of hereditary pancreatitis. Mutations in other genes, primarily serine protease inhibitor Kazal type 1 (SPINK1) and the cystic fibrosis transmembrane conductance regulator (CFTR) are also associated with hereditary pancreatitis. While chronic alcoholic pancreatitis may develop in the fourth or fifth decades, patients with hereditary pancreatitis may develop symptoms in the first or second decades of life. Hereditary pancreatitis is diagnosed either by detecting a causative gene mutation or by the presence of chronic pancreatitis in two first-degree or three second-degree relatives, in two or more generations, without precipitating factors and with a negative workup for known causes. Patients with hereditary pancreatitis may have recurrent acute pancreatitis and may develop pancreatic exocrine and endocrine insufficiency. Hereditary pancreatitis may involve premature trypsinogen activation or decreased control of trypsin. Recurrent inflammation can lead to acute pancreatitis and subsequently to chronic pancreatitis with parenchymal calcification. There is a markedly increased risk of pancreatic carcinoma compared with the general population. Patients are often referred for evaluation of pancreatitis, biliary or pancreatic ductal dilatation, jaundice, biliary obstruction, pancreatic duct stone or stricture, pancreatic pseudocysts, and for evaluation for malignancy. Medical treatment includes pancreatic enzyme supplementation, nutritional supplementation, diabetes management, and palliation of pain. Patients should avoid tobacco use and alcohol exposure. Hereditary pancreatitis is reviewed and recommendations for genetic testing are discussed.
chronic pancreatitis; cystic fibrosis; endoscopic surgical procedures; familial pancreatitis; hereditary pancreatitis; idiopathic pancreatitis; pancreatitis
AIM: To determine feasibility of liver transplantation in patients from the intensive care unit (ICU) by estimating graft and patient survival.
METHODS: This single center retrospective study included 39 patients who had their first liver transplant directly from the intensive care unit and 927 non-ICU patients who were transplanted from hospital ward or home between January 2005 and December 2010.
RESULTS: In comparison to non-ICU patients, ICU patients had a higher model for end-stage liver disease (MELD) at transplant (median: 37 vs 20, P < 0.001). Fourteen out of 39 patients (36%) required vasopressor support immediately prior to liver transplantation (LT) with 6 patients (15%) requiring both vasopressin and norepinephrine. Sixteen ICU patients (41%) were ventilator dependent immediately prior to LT with 9 patients undergoing percutaneous tracheostomy prior to transplantation. Twenty-five ICU patients (64%) required dialysis preoperatively. At 1, 3 and 5 years after LT, graft survival was 76%, 68% and 62% in ICU patients vs 90%, 81% and 75% in non-ICU patients. Patient survival at 1, 3 and 5 years after LT was 78%, 70% and 65% in ICU patients vs 94%, 85% and 79% in non-ICU patients. When formally comparing graft survival and patient survival between ICU and non-ICU patients using Cox proportional hazards regression models, both graft survival [relative risk (RR): 1.94, 95%CI: 1.09-3.48, P = 0.026] and patient survival (RR: 2.32, 95%CI: 1.26-4.27, P = 0.007) were lower in ICU patients vs non-ICU patients in single variable analysis. These findings were consistent in multivariable analysis. Although not statistically significant, graft survival was worse in both patients with cryptogenic cirrhosis (RR: 3.29, P = 0.056) and patients who received donor after cardiac death (DCD) grafts (RR: 3.38, P = 0.060). These findings reached statistical significance when considering patient survival, which was worse for patients with cryptogenic cirrhosis (RR: 3.97, P = 0.031) and patients who were transplanted with DCD livers (RR: 4.19, P = 0.033). Graft survival and patient survival were not significantly worse for patients on mechanical ventilation (RR: 0.91, P = 0.88 in graft loss; RR: 0.69, P = 0.56 in death) or patients on vasopressors (RR: 1.06, P = 0.93 in graft loss; RR: 1.24, P = 0.74 in death) immediately prior to LT. Trends toward lower graft survival and patient survival were observed for patients on dialysis immediately before LT, however these findings did not approach statistical significance (RR: 1.70, P = 0.43 in graft loss; RR: 1.46, P = 0.58 in death).
CONCLUSION: Although ICU patients when compared to non-ICU patients have lower survivals, outcomes are still acceptable. Pre-transplant ventilation, hemodialysis, and vasopressors were not associated with adverse outcomes.
Donor pool; Donor outcomes; Onor after cardiac death grafts; Liver graft survival; Patient survival
Inactivation and silencing of PTEN have been observed in multiple cancers, including follicular thyroid carcinoma. PTEN (phosphatase and tensin homologue deleted from chromosome 10) functions as a tumour suppressor by opposing the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) signalling pathway. Despite correlative data, how deregulated PTEN signalling leads to thyroid carcinogenesis is not known. Mice harbouring a dominant-negative mutant thyroid hormone receptor β (TRβPV/PV mice) spontaneously develop follicular thyroid carcinoma and distant metastases similar to human cancer. To elucidate the role of PTEN in thyroid carcinogenesis, we generated TRβPV/PV mice haploinsufficient for Pten (TRβPV/PVPten+/− mouse). PTEN deficiency accelerated the progression of thyroid tumour and increased the occurrence of metastasis spread to the lung in TRβPV/PVPten+/− mice, thereby significantly reducing their survival as compared with TRβPV/PVPten+/+ mice. AKT activation was further increased by two-fold in TRβPV/PVPten+/− mice thyroids, leading to increased activity of the downstream mammalian target of rapamycin (mTOR)–p70S6K signalling and decreased activity of the forkhead family member FOXO3a. Consistently, cyclin D1 expression was increased. Apoptosis was decreased as indicated by increased expression of nuclear factor-κB (NF-κB) and decreased caspase-3 activity in the thyroids of TRβPV/PVPten+/− mice. Our results indicate that PTEN deficiency resulted in increased cell proliferation and survival in the thyroids of TRβPV/PVPten+/− mice. Altogether, our study provides direct evidence to indicate that in vivo, PTEN is a critical regulator in the follicular thyroid cancer progression and invasiveness.
thyroid cancer; Pten; carcinogenesis; mouse model; mutations
Correlative data suggest that thyroid hormone receptor-β (TRβ) mutations could increase the risk of mammary tumor development, but unequivocal evidence is still lacking. To explore the role of TRβ mutants in vivo in breast tumor development and progression, we took advantage of a knock-in mouse model harboring a mutation in the Thrb gene encoding TRβ (ThrbPV mouse). Although in adult nulliparous females, a single ThrbPV allele did not contribute to mammary gland abnormalities, the presence of two ThrbPV alleles led to mammary hyperplasia in ~36% ThrbPV/PV mice. The ThrbPV mutation further markedly augmented the risk of mammary hyperplasia in a mouse model with high susceptibility to mammary tumors (Pten+/− mouse), as demonstrated by the occurrence of mammary hyperplasia in ~60% of Thrbpv/+Pten+/− and ~77% of ThrbPV/PV Pten+/− mice versus ~33% of Thrb+/+Pten+/− mice. The ThrbPV mutation increased the activity of signal transducer and activator of transcription (STAT5) to increase cell proliferation and the expression of the STAT5 target gene encoding β-casein in the mammary gland. We next sought to understand the molecular mechanism underlying STAT5 overactivation by TRβPV. Cell-based studies with a breast cancer cell line (T47D cells) showed that thyroid hormone (T3) repressed STAT5 signaling in TRβ-expressing cells through decreasing STAT5-mediated transcription activity and target gene expression, whereas sustained STAT5 signaling was observed in TRβPV-expressing cells. Collectively, these findings show for the first time that a TRβ mutation promotes the development of mammary hyperplasia via aberrant activation of STAT5, thereby conferring a fertile genetic ground for tumorigenesis.
thyroid hormone receptor; breast cancer; mammary tumors; Pten; mouse models
Aberrant expression and mutations of thyroid hormone receptor genes (TRs) are closely associated with several types of human cancers. To test the hypothesis that TRs could function as tumor suppressors, we took advantage of mice with deletion of all functional TRs (TRα1−/− TRβ−/−mice). As these mice aged, they spontaneously developed follicular thyroid carcinoma with pathological progression from hyperplasia to capsular invasion, vascular invasion, anaplasia and metastasis to the lung, similar to human thyroid cancer. Detailed molecular analysis revealed that known tumor promoters such as pituitary tumor-transforming gene were activated and tumor suppressors such as peroxisome proliferator-activated receptor γ and p53 were suppressed during carcinogenesis. In addition, consistent with the human cancer, AKT–mTOR–p70S6K signaling and vascular growth factor and its receptor were activated to facilitate tumor progression. This report presents in vivo evidence that functional loss of both TRα1 and TRβ genes promotes tumor development and metastasis. Thus, TRs could function as tumor suppressors in a mouse model of metastatic follicular thyroid cancer.
thyroid cancer; mouse model; mutations of thyroid hormone receptors
Accumulated evidence suggests that thyroid hormone receptor β (TRβ) could function as a tumor suppressor, but the detailed mechanisms by which TRβ inhibits tumorigenesis are not fully understood. The present studies explored the mechanisms by which TRβ acted to inhibit thyroid tumor development mediated by simian virus-40 (SV40). In mouse xenograft models, SV40 large T antigen (SV40Tag)-immortalized human thyroid epithelial (HTori) cells rapidly induced tumors, but the tumor development was totally blocked by TRβ stably expressed in HTori cells. Previous studies showed that the SV40Tag oncoprotein binds to and inactivates tumor suppressors p53 and retinoblastoma protein (Rb), thereby inducing tumorigenesis. Here we showed that one of the mechanisms by which TRβ suppressed tumor development was by competing with p53 and Rb for binding to SV40Tag. The interaction of TRβ with SV40Tag led to reactivation of Rb to inhibit cell cycle progression. TRβ- SV40Tag interaction also resulted in reactivating p53 to increase the expression of Pten, thus attenuating PI3K-AKT signaling to decrease cell proliferation and to induce apoptosis. The present study uncovered a novel action of TRβ as a tumor suppressor initiated via interfering with the recruitment of Rb and p53 by SV40Tag oncoprotein through protein-protein interaction, thereby acting to block tumor development.
Thyroid hormone receptor; tumor suppressor; tumorigenesis; thyroid hormone; xenograft models
Immune stimulation is a significant hurdle in the development of effective and safe RNA interference therapeutics. Here, we address this problem in the context of a mimic of microRNA-122 by employing novel nucleobase and known 2′-ribose modifications. The nucleobase modifications are analogues of adenosine and guanosine that contain cyclopentyl and propyl minor-groove projections. Via a site-by-site chemical modification analysis, we identify several immunostimulatory ‘hot spots’ within the miRNA guide strand at which single base modifications significantly reduce immune stimulation. A duplex containing one base modification on each strand proved to be most effective in preventing immune stimulation.
RNA interference; siRNA; microRNA; immunostimulation; nucleoside analogs
Natural orifice translumenal endoscopic surgery (NOTES) had its origins in numerous small animal studies primarily examining safety and feasibility. In human trials, safety and feasibility remain at the forefront; however, additional logistic, practical, and regulatory requirements must be addressed. The purpose of this paper is to evaluate and summarize published studies to date of NOTES in humans. The literature review was performed using PUBMED and MEDLINE databases. Articles published in human populations between 2007 and 2011 were evaluated. A review of this time period resulted in 48 studies describing procedures in 916 patients. Transcolonic and transvesicular procedures were excluded. The most common procedure was cholecystectomy (682, 75%). The most common approach was transvaginal (721, 79%). 424 procedures (46%) were pure NOTES and 491 (54%) were hybrid NOTES cases. 127 (14%) were performed in the United States of America and 789 (86%) were performed internationally. Since 2007, there has been major development in NOTES in human populations. A preponderance of published NOTES procedures were performed internationally. With further development, NOTES may make less invasive surgery available to a larger human population.
A randomized controlled field trial to evaluate the effectiveness of a single oral dose of 30 mg/kg of oxfendazole (OFZ) treatment for control of porcine cysticercosis was conducted in 4 rural villages of Angónia district, north-western Mozambique. Two hundred and sixteen piglets aged 4 months were selected and assigned randomly to OFZ treatment or control groups. Fifty-four piglets were treated at 4 months of age (T1), while another 54 piglets were treated at 9 months of age (T2) and these were matched with 108 control pigs from the same litters and raised under the same conditions. Baseline data were collected on the prevalence of porcine cysticercosis using antigen ELISA (Ag-ELISA), as well as knowledge and practices related to Taenia solium transmission based on questionnaire interviews and observations. All animals were followed and re-tested for porcine cysticercosis by Ag-ELISA at 9 and 12 months of age when the study was terminated. Overall prevalence at baseline was 5.1% with no significant difference between groups. At the end of the study, 66.7% of the controls were found positive, whereas 21.4% of the T1 and 9.1% of the T2 pigs were positive, respectively. Incidence rates of porcine cysticercosis were lower in treated pigs as compared to controls. Necropsy of 30 randomly selected animals revealed that viable cysts were present in none (0/8) of T2 pigs, 12.5% (1/8) of T1 pigs and 42.8% (6/14) of control pigs. There was a significant reduction in the risk of T. solium cysticercosis if pigs were treated with OFZ either at 4 months (OR = 0.14; 95% CI: 0.05–0.36) or at 9 months of age (OR = 0.05; 95% CI: 0.02–0.16). Strategic treatment of pigs in endemic areas should be further explored as a means to control T. solium cysticercosis/taeniosis.
Porcine cysticercosis is an infection of pigs caused by the larval stage of Taenia solium, a tapeworm that causes taeniosis in humans. The disease is very common in developing countries where it is a serious public health risk and causes significant economic losses in pig production. Many control strategies in developing countries have been of limited impact mainly due to poor socioeconomic and sanitary conditions. An effective treatment of infected pigs using inexpensive drugs may have potential as a long term control tool. We performed a randomized controlled trial to evaluate the effectiveness of oxfendazole treatment for control of porcine cysticercosis. We evaluated the prevalence and incidence of the disease in groups of pigs treated at 4 and 9 months of age and untreated pigs. We found that the prevalence and incidence of the disease in treated pigs was significantly lower than in untreated pigs. We conclude that treatment of pigs with oxfendazole in the last part of the fattening period is cost-effective in controlling porcine cysticercosis in endemic low-income areas but should be integrated with other control measures.
There is a high prevalence of Taenia solium taeniosis/cysticercosis in humans and pigs in the Eastern Cape Province (ECP) of South Africa. The objective of this study was to identify risk factors of porcine cysticercosis in select districts of the ECP. Data were collected in 2003 by interviewing 217 pig producers from the area. Blood samples were collected from 261 of their pigs, which were tested using two enzyme-linked immunosorbent assays (ELISA) for the presence of antibodies to cysticercosis. Frequencies of both owner- and pig-level characteristics were determined. For pig-level analysis, all bivariable and multivariable associations were determined using the surveylogistic procedure of the SAS/STAT® software to accommodate for the intraclass correlation that exists for clusters of pigs within one owner and for clusters of owners within a district. All tests for significance were performed at the α = 0.05 level, and adjusted odds ratios (aOR) and 95% confidence intervals (CI) were determined. Among the respondents, 48% of their households lacked a latrine, 98% slaughtered pigs at home, and 99% indicated that meat inspection services were not available. On bivariable analysis, there was a significant association between porcine infection and district (p = 0.003), breed (p = 0.041) and the absence of a latrine (p = 0.006). On multivariable analysis, the absence of a latrine was the only variable significantly associated with porcine infection (aOR = 1.89; 95% CI = 1.07, 3.35) (p = 0.028). The increased odds of porcine infection with households lacking a latrine contributes to our understanding of the transmission of this parasite in the ECP. Determining and addressing the risk factors for T. solium infection can potentially lower the very high prevalence in humans and pigs in this endemic area.
Systemic corticosteroids represent the standard treatment for autoimmune pancreatitis with IgG4-associated cholangitis. For steroid-dependent disease, azathioprine has been used for maintenance of remission. Mycophenolate mofetil has been used for transplant immunosuppression and more recently for autoimmune hepatitis; however, there are no case reports to date on the use of mycophenolate mofetil in adult patients with autoimmune pancreatitis. A patient with IgG4-mediated autoimmune pancreatitis and IgG4-associated cholangitis refractory to steroids and intolerant of azathioprine was treated with mycophenolate mofetil, which inhibits de novo guanosine synthesis and blockade of both B and T lymphocyte production. Introduction of mycophenolate mofetil and uptitration to 1000 mg by mouth twice daily over a treatment period of 4 mo was associated with improvement in the patient’s energy level and blood glucose control and was not associated with any adverse events. The patient was managed without a biliary stent. However, there was a return of symptoms, jaundice, increase in transaminases, and hyperbilirubinemia when the prednisone dose reached 11 mg per day. In the first report of mycophenolate mofetil use in an adult patient with IgG4-associated autoimmune pancreatitis and IgG4-associated cholangitis, the introduction of mycophenolate mofetil was safe and well-tolerated without adverse events, but it did not enable discontinuation of the steroids. Mycophenolate mofetil and other immunomodulatory therapies should continue to be studied for maintenance of remission in the large subset of patients with refractory or recurrent autoimmune pancreatitis.
Autoimmune diseases; Pancreatitis; Mycophenolate mofetil; Recurrence
Diagnostic tools appropriate for undertaking interventions to control helminth infections are key to their success. Many diagnostic tests for helminth infection have unsatisfactory performance characteristics and are not well suited for use in the parasite control programmes that are being increasingly implemented. Although the application of modern laboratory research techniques to improve diagnostics for helminth infection has resulted in some technical advances, uptake has not been uniform. Frequently, pilot or proof of concept studies of promising diagnostic technologies have not been followed by much needed product development, and in many settings diagnosis continues to rely on insensitive and unsatisfactory parasitological or serodiagnostic techniques. In contrast, PCR-based xenomonitoring of arthropod vectors, and use of parasite recombinant proteins as reagents for serodiagnostic tests, have resulted in critical advances in the control of specific helminth parasites. The Disease Reference Group on Helminths Infections (DRG4), established in 2009 by the Special Programme for Research and Training in Tropical Diseases (TDR) was given the mandate to review helminthiases research and identify research priorities and gaps. In this review, the diagnostic technologies relevant to control of helminth infections, either available or in development, are reviewed. Critical gaps are identified and opportunities to improve needed technologies are discussed.
While RNAi has traditionally relied on RNA duplexes, early evaluation of siRNAs demonstrated activity of the guide strand in the absence of the passenger strand. However, these single strands lacked the activity of duplex RNAs. Here, we report the systematic use of chemical modifications to optimize single-strand RNA (ssRNA)-mediated mRNA knockdown. We identify that 2′F ribose modifications coupled with 5′-end phosphorylation vastly improves ssRNA activity both in vitro and in vivo. The impact of specific chemical modifications on ssRNA activity implies an Ago-mediated mechanism but the hallmark mRNA cleavage sites were not observed which suggests ssRNA may operate through a mechanism beyond conventional Ago2 slicer activity. While currently less potent than duplex siRNAs, with additional chemical optimization and alternative routes of delivery, chemically modified ssRNAs could represent a powerful RNAi platform.
Current modifications used in small interfering RNAs (siRNAs), such as 2'-methoxy (2'-OMe) and 2'-fluoro (2'-F), improve stability, specificity or immunogenic properties but do not improve potency. These modifications were previously designed for use in antisense and not siRNA. We show, for the first time, that the siRNA-optimized novel 2'-O modifications, 2'-O-benzyl, and 2'-O-methyl-4-pyridine (2'-O-CH2Py(4)), are tolerated at multiple positions on the guide strand of siRNA sequences in vivo. 2'-O-benzyl and 2'-O-CH2Py(4) modifications were tested at each position individually along the guide strand in five sequences to determine positions that tolerated the modifications. The positions were combined together and found to increase potency and duration of siRNAs in vivo compared to their unmodified counterparts when delivered using lipid nanoparticles. For 2'-O-benzyl, four incorporations were tolerated with similar activity to the unmodified siRNA in vivo, while for 2'-O-CH2Py(4) six incorporations were tolerated. Increased in vivo activity was observed when the modifications were combined at positions 8 and 15 on the guide strand. Understanding the optimal placement of siRNA-optimized modifications needed for maximal in vivo activity is necessary for development of RNA-based therapeutics.
Vesicular stomatitis virus (VSV) is a potential oncolytic virus for treating glioblastoma multiforme (GBM), an aggressive brain tumor. Matrix (M) protein mutants of VSV have shown greater selectivity for killing GBM cells versus normal brain cells than VSV with wild-type M protein. The goal of this research was to determine the contribution of death receptor and mitochondrial pathways to apoptosis induced by an M protein mutant (M51R) VSV in U87 human GBM tumor cells. Compared to controls, U87 cells expressing a dominant negative form of Fas (dnFas) or overexpressing Bcl-XL had reduced caspase-3 activation following infection with M51R VSV, indicating that both the death receptor pathway and mitochondrial pathways are important for M51R VSV-induced apoptosis. Death receptor signaling has been classified as type I or type II, depending on whether signaling is independent (type I) or dependent on the mitochondrial pathway (type II). Bcl-XL overexpression inhibited caspase activation in response to a Fas-inducing antibody, similar to the inhibition in response to M51R VSV infection, indicating that U87 cells behave as type II cells. Inhibition of apoptosis in vitro delayed, but did not prevent, virus-induced cell death. Murine xenografts of U87 cells that overexpress Bcl-XL regressed with a time course similar to that of control cells following treatment with M51R VSV, and tumors were not detectable at 21 days postinoculation. Immunohistochemical analysis demonstrated similar levels of viral antigen expression but reduced activation of caspase-3 following virus treatment of Bcl-XL-overexpressing tumors compared to controls. Further, the pathological changes in tumors following treatment with virus were quite different in the presence versus the absence of Bcl-XL overexpression. These results demonstrate that M51R VSV efficiently induces oncolysis in GBM tumor cells despite deregulation of apoptotic pathways, underscoring its potential use as a treatment for GBM.
HER2 overexpression stimulates cell growth in p53-mutated cells while it inhibits cell proliferation in those with wild-type p53, but the molecular mechanism is unknown. The Dmp1 promoter was activated by HER2/neu through the PI3K-Akt-NF-κB pathway, which in turn stimulated Arf transcription. Binding of p65 and p52 subunits of NF-κB was demonstrated to the Dmp1 promoter and that of Dmp1 to the Arf promoter upon HER2/neu overexpression. Both Dmp1 and p53 were induced in pre-malignant lesions from MMTV-neu mice and mammary tumorigenesis was significantly accelerated in both Dmp1+/− and Dmp1−/− mice. Selective deletion of Dmp1 and/or overexpression of Tbx2/Pokemon was found in >50 % of wild-type HER2/neu carcinomas while the involvement of Arf, Mdm2, or p53 was rare. Tumors from Dmp1+/−, Dmp1−/−, and wild-type neu mice with hemizygous Dmp1 deletion showed significant downregulation of Arf and p21Cip1/WAF1, showing p53 inactivity and more aggressive phenotypes than tumors without Dmp1 deletion. Notably, endogenous hDMP1 mRNA decreased when HER2 was depleted in human breast cancer cells. Our study demonstrates the pivotal roles of Dmp1 in HER2/neu-p53 signaling and breast carcinogenesis.
Dmp1 (Dmtf1); HER2/neu; NF-κB; Arf; p53; Tbx2; Pokemon; breast cancer
Drosophila melanogaster is one of the most well studied genetic model organisms, nonetheless its genome still contains unannotated coding and non-coding genes, transcripts, exons, and RNA editing sites. Full discovery and annotation are prerequisites for understanding how the regulation of transcription, splicing, and RNA editing directs development of this complex organism. We used RNA-Seq, tiling microarrays, and cDNA sequencing to explore the transcriptome in 30 distinct developmental stages. We identified 111,195 new elements, including thousands of genes, coding and non-coding transcripts, exons, splicing and editing events and inferred protein isoforms that previously eluded discovery using established experimental, prediction and conservation-based approaches. Together, these data substantially expand the number of known transcribed elements in the Drosophila genome and provide a high-resolution view of transcriptome dynamics throughout development.
Ferritin binds specifically and saturably to a variety of cell types, and recently several ferritin receptors have been cloned. TIM-2 is a specific receptor for H ferritin (HFt) in the mouse. TIM-2 is a member of the T cell immunoglobulin and mucin domain containing (TIM) protein family and plays an important role in immunity. The expression of TIM-2 outside of the immune system indicates that this receptor may have broader roles. We tested whether ferritin binding to TIM-2 can serve as an iron delivery mechanism. TIM-2 was transfected into normal (TCMK-1) mouse kidney cells, where it was appropriately expressed on the cell surface. HFt was labeled with 55Fe and 55Fe-HFt was incubated with TIM-2 positive cells or controls. 55Fe-HFt uptake was observed only in TIM-2 positive cells. HFt uptake was also seen in A20 B cells, which express endogenous TIM-2. TIM-2 levels were not increased by iron chelation. Uptake of 55Fe-HFt was specific and temperature-dependent. HFt taken up by TIM-2 positive cells transited through the endosome and eventually entered a lysosomal compartment, distinguishing the HFt pathway from that of transferrin, the classical vehicle for cellular iron delivery. Iron delivered following binding of HFt to TIM-2 entered the cytosol and became metabolically available, resulting in increased levels of endogenous intracellular ferritin. We conclude that TIM-2 can function as an iron uptake pathway.
In this study, we pilot tested an in vitro assay of cancer killing activity (CKA) in circulating leukocytes of 22 cancer cases and 25 healthy controls.
Using a human cervical cancer cell line, HeLa, as target cells, we compared the CKA in circulating leukocytes, as effector cells, of cancer cases and controls. The CKA was normalized as percentages of total target cells during selected periods of incubation time and at selected effector/target cell ratios in comparison to no-effector-cell controls.
Our results showed that CKA similar to that of our previous study of SR/CR mice was present in human circulating leukocytes but at profoundly different levels in individuals. Overall, males have a significantly higher CKA than females. The CKA levels in cancer cases were lower than that in healthy controls (mean ± SD: 36.97 ± 21.39 vs. 46.28 ± 27.22). Below-median CKA was significantly associated with case status (odds ratio = 4.36; 95% Confidence Interval = 1.06, 17.88) after adjustment of gender and race.
In freshly isolated human leukocytes, we were able to detect an apparent CKA in a similar manner to that of cancer-resistant SR/CR mice. The finding of CKA at lower levels in cancer patients suggests the possibility that it may be of a consequence of genetic, physiological, or pathological conditions, pending future studies with larger sample size.
Selective drugs targeting dysregulated oncogenic pathways are promising cancer therapies. Because the mammalian target of rapamycin complex 1 (mTORC1) pathway is hyperactivated in human follicular thyroid cancer (FTC), we hypothesized that its inhibition could block cancer development and progression. We, therefore, analyzed the effect of a treatment with a specific mTORC1 inhibitor (RAD001) in a faithful mouse model of FTC with constitutive mTORC1 activation (TRβPV/PVPten+/− mice). The treatment did not prevent capsular and vascular invasion of the thyroid and the occurrence of lung metastasis. However, it substantially decelerated thyroid tumor growth, thereby prolonging TRβPV/PVPten+/− mouse life span. RAD001 efficiently inhibited mTORC1 activity, as shown by the reduced phosphorylation of its downstream targets involved in the activity of the translation machinery, such as ribosomal S6 kinase (p70S6K), eukaryotic translation initiation factor 4E binding protein (4E-BP1) and the eukaryotic translation initiation factors eIF-4B and eIF-4G. Whereas mTORC1 signaling inhibition did not alter cell apoptosis, it induced a significant decrease in cell proliferation that was associated with the reduced abundance and altered activity of key regulators of cell cycle progression. Altogether, our data indicate that mTORC1 signaling plays a major role in the integration of the mitogenic signal in FTC. Therefore, our preclinical study with a relevant mouse model of FTC demonstrates for the first time that RAD001 efficaciously stabilizes cancer growth although it does not prevent its fatal outcome. In conclusion, our work underscores that in the treatment of FTC patients, RAD001 can only be used in combination with drugs and therapies inducing tumor shrinkage and blocking metastasis.
Despite growing awareness of the importance of controlling neglected tropical diseases as a contribution to poverty alleviation and achieving the Millennium Development Goals, there is a need to up-scale programmes to achieve wider public health benefits. This implementation deficit is attributable to several factors but one often overlooked is the specific difficulty in tackling diseases that involve both people and animals - the zoonoses. A Disease Reference Group on Zoonoses and Marginalised Infectious Diseases (DRG6) was convened by the Special Programme for Research and Training in Tropical Diseases (TDR), a programme executed by the World Health Organization and co-sponsored by UNICEF, UNDP, the World Bank and WHO. The key considerations included: (a) the general lack of reliable quantitative data on their public health burden; (b) the need to evaluate livestock production losses and their additional impacts on health and poverty; (c) the relevance of cross-sectoral issues essential to designing and implementing public health interventions for zoonotic diseases; and (d) identifying priority areas for research and interventions to harness resources most effectively. Beyond disease specific research issues, a set of common macro-priorities and interventions were identified which, if implemented through a more integrated approach by countries, would have a significant impact on human health of the most marginalised populations characteristically dependent on livestock.
Mice may be useful for studies of skeletal aging, but there are limited data on changes in bone structure and strength over their lifespan. We obtained bones from female and male BALB/c mice at ages 2, 4, 7, 12 and 20 months and evaluated their structural, densitometric and mechanical properties. MicroCT of the mid-diaphysis of the femur and radius indicated that during skeletal growth (2–7 months) bone cross-sectional size (area, moment of inertia) increased rapidly; during aging (7–20 months) cortical area was maintained while moment of inertia continued to increase. Bones from females were smaller than males at young ages, but not at later ages. Changes in whole-bone stiffness and strength reflected the changes in bone size, with a rapid increase from 2–7 months followed by little or no change. In contrast, energy-to-fracture declined with aging. Cortical tissue mineral density (TMD) increased during growth and was maintained with aging. MicroCT of trabecular bone revealed age-related changes that were site-dependent. The proximal tibia showed a clear pattern of age-related decline in trabecular BV/TV, with progressive decreases after 4 months in both sexes; lumbar (L5) vertebra had more modest age-related declines; in contrast, caudal (Ca7) vertebra had increasing BV/TV with aging. Overall, we found no evidence that females had more pronounced age-related deterioration than males. We conclude that bones from aging female and male BALB/c mice exhibit many of the changes seen in humans, and are therefore a clinically relevant model for studies of skeletal aging.
bone structure; bone strength; bone mineralization; aging; mouse model